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Living Phanerochaete chrysosporium mycelia were used to remove heavy metals of Cd(II), Cu(II) and Zn(II) in auqeous solution. The uptake of heavy metal by the mycelia was dependent on the environmental conditions. The optimum biosorption conditions of Cd(II), Cu(II), and Zn(II) were pH 5.5–6.5 at 37◦C, and 6 h. Under these conditions, the fungal biosorbent removed Cd(II), Cu(II), and Zn(II) rapidly and efficiently with maximum metal removal capacities of 59.77 mg/g, 74.78 mg/g, and 54.12 mg/g, respectively. The pseudo second-order kinetic model was superior to the pseudo first-order kinetic model, which indicated that the change of the surface sorption sites number was proportional to the square of the remaining unoccupied surface sites number. Fourier transform infrared (FT-IR) spectra indicated that hydroxyl and carboxyl groups were relevant to biosorption. Some crystal particles were found on the surface of the P. chrysosporium under scanning electron microscopy. Energy dispersive X-ray analysis and FT-IR revealed that amino acids and proteins were involved in binding metal ions. The results demonstrated that P. chrysosporium was a good potential biosorbent for adsorbing heavy metals. [ABSTRACT FROM AUTHOR]

Purpose: The suboptimal features of the spectral properties of the leading fluorescence resonance energy transfer (FRET) pair, cyan fluorescent protein (CFP)/yellow fluorescent protein (YFP), limit the full promise of FRET imaging. To overcome the drawbacks, we developed FRET-based, intra-molecular biosensors consisting of CFP/discomona sp red fluorescent protein (DsRed) or green fluorescent protein (GFP)/DsRed as donor/acceptor fluorophores.Procedures: The biosensors were expressed in NIH3T3 cells. In vitro fluorescence spectroscopy and Rho GTPase activation assays were used to confirm that Rac1 or Cdc42 was activated in serum-starved cells following stimulation with insulin or bradykinin. The transient changes of the amount, location, and translocation of activated Rac1 or Cdc42 in living cells were tracked with confocal imaging.Results: The increase of FRET efficiency was achieved in the cells expressing the biosensors and was proportional to the levels of activated Rac1 or Cdc42. The localized, transitional, and transient FRET signals were directly and quantitatively imaged with high spatial and temporal resolution. The biosensors were used to analyze and judge the GEF or GAP activities of putative regulatory proteins for Rac1 or Cdc42.Conclusion: DsRed is a more suitable acceptor in FRET pair with CFP than with GFP in terms of the spectral overlap between the donor and acceptor. The approach can also be applied to many other types of protein behavior in living cells. [ABSTRACT FROM AUTHOR]

Potassium channel tetramerization domain-containing 10 gene ( KCTD10) belongs to the polymerase delta-interacting protein 1 ( PDIP1) gene family. Mouse KCTD10 was thought to interact with proliferating cell nuclear antigen and the small subunit of polymerase δ, and to have roles in DNA repair, DNA replication and cell-cycle control. To better understand the regulatory mechanism of KCTD10 expression, we characterized the promoter of human KCTD10 containing a 639 bp fragment of the 5′-flanking region (−609/+30). A primer extension assay identified the transcription start site as an A, 63 bp upstream of the start codon. The promoter region contains neither a TATA box nor a CCAAT box, but a CpG island was found near to the transcription start site. Deletion mutagenesis showed that the region from −108 to +30 was indispensable to the promoter activity, and site-directed mutation analysis in this proximal promoter region of KCTD10 revealed two important transcription regulatory elements of specificity protein 1 (SP1) and activating protein-2 (AP-2). An in vivo chromatin immunoprecipitation assay further demonstrated that SP1 and AP-2α could bind to this proximal promoter region. Moreover, using a luciferase reporter assay, quantitative real-time RT-PCR and western blot analysis, both overexpression and RNA interference of SP1 and AP-2α indicated that the binding of SP1 to the proximal promoter region stimulated the promoter activity and endogenous KCTD10 expression, whereas binding of AP-2α to this region showed opposite effects. [ABSTRACT FROM AUTHOR]

Background: Activator protein 2 gamma (AP-2gamma) is a member of the transcription factor activator protein-2 (AP-2) family, which is developmentally regulated and plays a role in human neoplasia. AP-2gamma has been found to be overexpressed in most breast cancers, and have a dual role to inhibit tumor initiation and promote tumor progression afterwards during mammary tumorigensis.Methods: To identify the gene targets that mediate its effects, we performed chromatin immunoprecipitation (ChIP) to isolate AP-2gamma binding sites on genomic DNA from human breast cancer cell line MDA-MB-453.Results: 20 novel DNA fragments proximal to potential AP-2gamma targets were obtained. They are categorized into functional groups of carcinogenesis, metabolism and others. A combination of sequence analysis, reporter gene assays, quantitative real-time PCR, electrophoretic gel mobility shift assays and immunoblot analysis further confirmed the four AP-2gamma target genes in carcinogenesis group: ErbB2, CDH2, HPSE and IGSF11. Our results were consistent with the previous reports that ErbB2 was the target gene of AP-2gamma. Decreased expression and overexpression of AP-2gamma in human breast cancer cells significantly altered the expression of these four genes, indicating that AP-2gamma directly regulates them.Conclusion: This suggested that AP-2gamma can coordinate the expression of a network of genes, involving in carcinogenesis, especially in breast cancer. They could serve as therapeutic targets against breast cancers in the future. [ABSTRACT FROM AUTHOR]